VITAMIN K, METABOLISM AND FUNCTION

维生素 K、新陈代谢和功能

• 批准号: 2901372
• 负责人: REIDAR WALLIN
• 金额: $ 22.58万
• 依托单位: WAKE FOREST UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-04-01 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2901372
• 关键词: Sf9 cell line; carboxylation; chemical binding; chemical models; coagulation factor IX; coagulation factor VII; cofactor; decarboxylases; drug resistance; enzyme complex; enzyme reconstitution; glutathione transferase; laboratory rabbit; laboratory rat; molecular site; oxidoreductase; pharmacokinetics; polymerase chain reaction; posttranslational modifications; protein biosynthesis; protein sequence; protein structure function; vitamin K; vitamin metabolism; warfarin

DESCRIPTION: Understanding vitamin K-dependent proteins will require insight into their biosynthesis and regulation. Unique for these proteins is their vitamin K-dependent post-translational modification. The modification is carried out by the _-carboxylase, an integral protein of the ER membrane which uses the reduced form of vitamin K (vitamin KH2) as cofactor. The reduced cofactor is produced by the enzyme Vitamin K 3,4-Epoxide reductase (VKOR) which is the target for the anticoagulant drug warfarin. Despite numerous attempts to purify VKOR, the molecular components that constitute the enzyme have not been identified. The applicants have been able to show that VKOR is an enzyme complex in the ER membrane and they propose that assembly of the complex is similar to assembly of the lipoxygenase complex on the nuclear envelope membrane. Both enzymes incorporates a member of the glutathione-S-transferase gene family as part of the lipid-protein enzyme complex. Experiments are proposed to complete the model of the VKOR enzyme complex and unveil the basis for genetic resistance to warfarin. Recombinant VKOR will be used to increase the capacity of cell lines to produce functional clotting factors VII and IX. It is our hypothesis that vitamin KH2 cofactor production by VKOR is a limiting factor in cellular production of these recombinant proteins. Although 8-10 different vitamin K-dependent proteins have been shown to be made extrahepatically only some have been identified. They include the coagulation factors prothrombin and protein S, the growth arrest specific gene 6 product Gas6 and the bone and cartilage resident proteins, osteocalsin and matrix GLA protein (MGP). In the last specific aim of this application experiments are proposed to identify new vitamin K-dependent proteins by vitamin K-dependent radioactive labeling. The proposed experiments will provide a better understanding of vitamin K metabolism and vitamin K function and have an impact on prophylactic medicine concerned with recombinant vitamin K-dependent coagulation factors and warfarin anticoagulation. The experiments should also contribute to the exciting, rapidly expanding area of research on extra-hepatic vitamin K-dependent proteins.

描述：理解依赖维生素K的蛋白质需要 洞察它们的生物合成和调控。这些蛋白质是独一无二的 是它们依赖维生素K的翻译后修饰。这个 修饰是由_-羧基酶进行的，_-羧化酶是 使用还原形式的维生素K(维生素Kh2)作为 辅因。还原的辅因子是由维生素K酶产生的 3，4-环氧化物还原酶(VKOR)是抗凝血药物的靶点 华法林。尽管多次尝试提纯VKOR，但分子 构成这种酶的成分尚未确定。这个 申请者已经能够证明VKOR是内质网中的一种酶复合体 膜，他们提出复合体的组装类似于 脂氧合酶复合体在核膜上的组装。两者都有 酶结合了谷胱甘肽-S-转移酶基因家族的一个成员 作为脂肪-蛋白质酶复合体的一部分。建议进行实验，以 完成VKOR酶复合体的模型并揭示其基础 对华法林的遗传抗性。将使用重组VKOR来增加 细胞系产生功能性凝血因子VII和 IX.我们的假设是VKOR产生的维生素Kh2辅因子是一个 这些重组蛋白在细胞生产中的限制因素。 尽管8-10种不同的维生素K依赖蛋白已被证明是 肝外发生的只有一些已经被鉴定出来。它们包括 凝血因子凝血酶原和蛋白S对生长停滞的特异性 基因6产物Gas6以及骨骼和软骨驻留蛋白， 骨钙素和基质GLA蛋白(MGP)。在此的最后一个具体目标中 建议进行应用实验以确定新的维生素K依赖 蛋白质的维生素K依赖的放射性标记。建议数 实验将提供更好的了解维生素K代谢和 维生素K的作用及其对预防医学的影响 重组维生素K依赖凝血因子和华法林 抗凝剂。这些实验也应该有助于激动人心的， 迅速扩大的肝外维生素K依赖研究领域 蛋白质。